DNA microarrays (also referred to as DNA chips) are used as a powerful sensor for detecting biomolecules such as DNA or RNA in samples derived from living bodies. By using DNA microarrays, it is possible to carry out detection or sequencing which requires hundreds to tens of thousands of times of the detection or sequencing operations simultaneously at a time. DNA microarrays have hundreds to tens of thousands of detection points (spots) aligned in rows on a several square centimeters to ten and several square centimeters support made of a glass slide or silicon chip. At each detection point, a single-stranded nucleic acid polymer (gene fragment) with one known base sequence is attached to the support. In other words, DNA microarray is a miniature array of a large number of probe polymers with different base sequences. By applying an aqueous solution of a nucleic acid sample labeled with a fluorophore to such a DNA microarray, only nucleic acid polymers with complementary base sequence hybridize to probes. The DNA microarray is then washed, and only the target nucleic acid polymers hybridized to the probes remain on the DNA microarray. By illuminating with excitation light, fluorescence light is emitted from the fluorophore in the target nucleic acid polymers remaining on the DNA microarray. It can be determined whether there are target nucleic acid polymers present in the nucleic acid sample by detecting the fluorescence light.
DNA microarrays can be divided roughly into the two types according to the fabrication methods: photolithography type and spotting type.
The photolithography-type DNA microarrays are made by synthesizing a large number of DNA (oligonucleotides) with desired different base sequences on a support (chip or sheet) by the photolithography technology used in the fabrication of semiconductor integrated circuits. DNA microarrays with high-density DNA detection points are already put to practical use (U.S. Pat. Nos. 5,744,305 and 5,445,934).
On the other hand, the spotting-type microarrays are made by placing drops containing DNA prepared beforehand one by one on a solid support (U.S. Pat. No. 5,807,522).
The two types of DNA microarrays described above have the following different features.
The photolithography-type DNA microarrays have the advantage of a high measurement sensitivity and its assured reproducibility and being usable for the SNP (Single Nucleotide Polymorphism) analysis because DNA detection points can be made very small and DNA can be grown uniformly. For the fabrication of the photolithography-type of DNA microarrays, an expensive semiconductor manufacturing equipment (“stepper”) that costs several million dollars is needed. In addition, the labor and cost for preparing a large number of photomasks increases with the increase of the number of DNA synthesized. The photolithography-type DNA microarrays is therefore very expensive and used only by some research laboratories that have ample research funds as those of pharmaceutical companies at the present time. It is also practically impossible at present to fabricate the photolithography-type DNA microarrays one by one according to the requirements of individual researchers.
The spotting-type microarrays is fabricated by placing drops containing DNA probes on a solid support and drying up, and hence there is inherently a limit to the reduction of the size of DNA detection points. Further, the density and uniformity of the DNA probes attached to the support are not assured so much as they are for the photolithography-type DNA microarrays. This is one of the fundamental factors that prevent the standardization of the spotting-type microarrays. For the fabrication of the spotting-type of DNA microarrays, an expensive, large-scale equipment that costs several tens of millions of dollars is also needed, though not expensive as the equipment for the photolithography-type of DNA microarrays.